Abstract
Synthetic materials are integral components of consumables and durable goods and indispensable in our modern world. Polyesters are the most versatile bulk- and specialty-polymers, but their production is not sustainable, and their fate at end-of-life of great concern. Bioplastics are highly regarded alternatives but have shortcomings in material properties and commercial competitiveness with conventional synthetic plastics. These constraints have limited the success in global markets. Enabling bio-production of advanced bioplastics with superior properties from waste-derived feedstocks could change this.
We have created microbial cell factories that can produce a range of aliphatic and aromatic polyesters. A ΔphaC1 mutant of Cupriavidus necator H16 was complemented with hydroxyacyl-CoA transferases from either Clostridium propionicum (pct540) or Clostridium difficile (hadA), respectively. These were combined with a mutant PHA synthase (phaC1437) from Pseudomonas sp. MBEL 6-19, which rescued the PHA− phenotype of the knock-out mutant and allowed polymerization of various hydroxy carboxylates, including phloretic acid. This is the first-time, incorporation of an aromatic ring in the backbone of a biological polyester was achieved. Polymers contain para-hydroxyphenyl subunits are structurally analogous to synthetic aromatic polyesters like PET and high-strength polyarylates.
In a further advance, the transgenic strain was cultivated in a bio-electrochemical system under autotrophic conditions, enabling synthesis of aromatic bio-polyesters from H2 and O2 generated in situ, while assimilating CO2. Follow-up elementary flux-mode analysis established the feasibility of de novo production of twenty different polyesters from five different carbon- and energy-sources. This comprehensive study opens the door to sustainable bio-production of high-performance thermoplastics and thermosets.
Significance statement New biomaterials can facilitate transition to a carbon-neutral chemical industry and a circular economy while at the same time preventing accumulation of plastic wastes in the environment. This can be accomplished by developing “drop-in” replacements for existing fossil carbon-based plastics. To that end, this work demonstrates for the first time that biocatalytic polymerization can incorporate an aromatic para-hydroxyphenyl carbonic acid into the backbone of a bio-polyester. This was accomplished using a genetically engineered microbial cell factory that assimilates carbon dioxide using hydrogen that is produced electrochemically in situ. A bio-electrochemical system eliminates the need for external supply of oxyhydrogen and avoids explosive mixtures, opening the door for sustainable production of biomaterials analogous to aromatic bulk-polyesters such as PET and high-performance “liquid-crystal polymers”.
Highlights
- Biocatalytic formation of aromatic polyesters with structural analogy to polyarylates
- Production of novel PHAs from CO2 and H2+O2 produced in situ in a bio-electrochemical system
- Expression-level of PHA synthase and molecular weight of polyesters are inversely correlated
- In silico design and analysis of pathways towards novel PHAs from different carbon-sources
Competing Interest Statement
The authors have declared no competing interest.
Abbreviations
- 3HP
- 3-hydroxypropionic acid
- 4HB
- 4-hydroxybutyric acid
- 6HC
- 6-hydroxycaproic acid
- BES
- bio-electrochemical system
- BM
- biomass
- CDW
- cell dry weight
- DAP
- diaminopimelate
- DSC
- differential scanning calorimetry
- EMA
- elementary flux-mode analysis
- GC-MS
- gas chromatography–mass spectrometry
- GPC
- gel permeation chromatography
- HA
- hydroxy acid
- MA
- mandelic acid
- MES
- microbial electrosynthesis
- Mn
- number-average molecular weight
- MSM
- mineral salts medium
- MW
- weight-average molecular weight
- NMR
- nuclear magnetic resonance
- OD
- optical density
- PA
- phloretic acid
- PDI
- polydispersity index
- PET
- polyethylene terephthalate
- PHA
- polyhydroxyalkanoate
- PHB
- polyhydroxybutyrate
- PHBV
- polyhydroxybutyrate-co-hydroxyvalerate
- PheLA
- phenyllactic acid
- PLA
- polylactic acid
- RB
- rich broth
- rpm
- rounds per minute
- VFA
- volatile fatty acid